Kovacs, G. G., Ferrer, I., Grinberg, L. T., Alafuzoff, I., Attems, J., Budka,H., ... Dickson, D. W. (2016). Aging-related tau astrogliopathy (ARTAG):harmonized evaluation strategy. Acta Neuropathologica, 131(1), 87-102.DOI: 10.1007/s00401-015-1509-x

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Aging-related tau astrogliopathy (ARTAG): harmonized evaluation strategy Gabor G. Kovacs1, Isidro Ferrer2, Lea T. Grinberg3,4, Irina Alafuzoff5, Johannes Attems6, Herbert Budka7, Nigel J. Cairns8, John F. Crary9, Charles Duyckaerts10, Bernardino Ghetti11, Glenda M. Halliday12, James W. Ironside13, Seth Love14, Ian R. Mackenzie15, David G. Munoz16, Melissa E. Murray17, Peter T. Nelson18, Hitoshi Takahashi19, John Q. Trojanowski20, Olaf Ansorge21, Thomas Arzberger22, Atik Baborie23, Thomas G. Beach24, Kevin F. Bieniek17, Eileen H. Bigio25, Istvan Bodi26, Brittany N. Dugger24,27, Mel Feany28, Ellen Gelpi29, Stephen M. Gentleman30, Giorgio Giaccone31, Kimmo J. Hatanpaa32, Richard Heale6, Patrick R. Hof33, Monika Hofer21, Tibor Hortobágyi34, Kurt Jellinger35, Gregory A. Jicha36, Paul Ince37, Julia Kofler38, Enikö Kövari39, Jillian J. Kril40, David M. Mann41, Radoslav Matej42, Ann C. McKee43, Catriona McLean44, Ivan Milenkovic1, 45, Thomas J. Montine46, Shigeo Murayama47, Edward B. Lee20, Jasmin Rahimi1, Roberta D. Rodriguez48, Annemieke Rozemüller49, Julie A. Schneider50, Christian Schultz51, William Seeley3, Danielle Seilhean10, Colin Smith13, Fabrizio Tagliavini31, Masaki Takao52, Dietmar Rudolf Thal53,54, Jon B. Toledo20, Markus Tolnay55, Juan C. Troncoso56, Harry V. Vinters57, Serge Weis58, Stephen B. Wharton37, Charles L. White, III32, Thomas Wisniewski59, John M. Woulfe60, Masahito Yamada61, and Dennis W. Dickson17 Affiliations: 1: Institute of Neurology, Medical University of Vienna, Vienna, Austria; 2: Institute of Neuropathology, Bellvitge University Hospital, University of Barcelona, Hospitalet de Llobregat, Barcelona, Spain; 3: Memory and Aging Center, Department of Neurology, University of California, San Francisco 4: Department of Pathology, LIM-22, University of Sao Paulo Medical School, Sao Paulo, Brazil 5: Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden 6: Institute of Neuroscience, Newcastle University, Newcastle upon Tyne, UK 7: Institute of Neuropathology, University Hospital Zürich, Switzerland 8: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA 9: Department of Pathology, Fishberg Department of Neuroscience, Friedman Brain Institute, and the Ronald M. Loeb Center for Alzheimer’s Disease, Icahn School of Medicine at Mount Sinai 10: Neuropathology Department, Hopital de La Salpetrière, AP-HP, UPMC-Sorbonne-University, Paris, France 11: Indiana University School of Medicine Department of Pathology and Laboratory Medicine, Indianapolis, IN, USA 12: GMH - Neuroscience Research Australia and the University of New South Wales, Sydney, NSW, Australia 13: Centre for Clinical Brain Sciences, University of Edinburgh, Edinburgh UK 14: Institute of Clinical Neurosciences, University of Bristol, Learning & Research level 2, Southmead Hospital, Bristol, UK 15: Department of Pathology and Laboratory Medicine, University of British Columbia,Vancouver, Canada 16: Division of Pathology, St. Michael’s Hospital 30 Bond St, Toronto, ON, Canada 17: Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL 32224, USA

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18: Department of Pathology and Sanders-Brown Center on Aging, University of Kentucky, Lexington, KY USA 40536, USA 19: Department of Pathology, Brain Research Institute, Niigata University, Niigata 951-8585, Japan 20: Center for Neurodegenerative Disease Research, Institute on Aging and Department of Pathology & Laboratory Medicine of the Perelman School of Medicine at the University of Pennsylvania 21: Department of Neuropathology, John Radcliffe Hospital, Oxford, UK 22: Department of Psychiatry and Psychotherapy, and Centre for Neuropathology and Prion Research, Ludwig-Maximilians-University Munich, Germany 23: Department of Neuropathology, Walton Centre, Liverpool, UK 24: Civin Laboratory for Neuropathology, Banner Sun Health Research Institute, Sun City, AZ 85351, USA 25: Northwestern ADC Neuropathology Core, Northwestern University Feinberg School of Medicine, Chicago, IL, USA 26: Clinical Neuropathology, King’s College Hospital and London Neurodegenerative Brain Bank, London, UK 27: University of California San Francisco, Institute for Neurodegenerative Diseases, San Francisco, CA, USA 28: Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA 29: Neurological Tissue Bank of the Biobank-Hospital Clinic-IDIBAPS, Institut d’Investigacions Biomediques August Pi i Sunyer, Barcelona, Spain 30: Department of Medicine, Imperial College London, London, UK 31: IRCCS Foundation “Carlo Besta” Neurological Institute, Milan, Italy 32: Department of Pathology, University of Texas Southwestern Medical Center, Dallas, TX, USA 33: Fishberg Department of Neuroscience, Friedman Brain Institute, and Ronald M. Loeb Center for Alzheimer’s Disease, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA 34: Department of Neuropathology, Institute of Pathology, Faculty of Medicine, University of Debrecen, Nagyerdei krt. 98, H-4032, Debrecen, Hungary. 35: Institute of Clinical Neurobiology; Alberichgasse 5/13; A-1150 Vienna 36: Department of Neurology and Sanders-Brown Center on Aging, University of Kentucky, Lexington, KY 40536, USA 37: Sheffield Institute for Translational Neuroscience, University of Sheffield, UK 38: Department of Pathology, University of Pittsburgh, Pittsburgh, PA, USA 39: Department of Mental Health and Psychiatry, University Hospitals and University of Geneva School of Medicine, Geneva, Switzerland 40: Discipline of Pathology, Sydney Medical School, The University of Sydney, Sydney NSW 2006, Australia 41: Institute of Brain, Behaviour and Mental Health, Manchester University Faculty of Medical and Health Sciences, Manchester, UK 42: Department of Pathology and Molecular Medicine, Thomayer Hospital, Prague 4, Czech Republic 43: Department of Neurology and Pathology, Boston University School of Medicine and VA Healthcare system, Boston, MA 02118, USA 44: Department of Anatomical Pathology, Alfred Hospital, Prahran, Victoria, 3004, Australia 45: Department of Neurology, Medical University of Vienna, Vienna, Austria 46: Department of Pathology, University of Washington, Seattle, WA, USA 47: Department of Neuropathology (the Brain Bank for Aging Research), Tokyo Metropolitan

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Geriatric Hospital & Institute of Gerontology, Tokyo, Japan 48: Physiopathology in Aging Lab/Brazilian Aging Brain Study Group-LIM22, University of Sao Paulo Medical School, Sao Paulo, Brazil 49: Netherlands Brainbank and Dept. of Pathology, VU University Medical Center, Amsterdam, The Netherlands 50: Departments of Pathology and Neurological Sciences, Rush University Medical Center, Chicago IL, USA 51: Institute of Neuroanatomy, Centre for Biomedicine and Medical Technology Mannheim, Medical Faculty Mannheim, Heidelberg University, Germany 52: Department of Neurology, Saitama Medical University International Medical Center, Saitama, Japan 53: Institute of Pathology – Laboratory of Neuropathology, University of Ulm, D-89081 Ulm, Germany 54: Department of Neuroscience, KU-Leuven, B-3000 Leuven, Belgium 55: Institute of Pathology, University Hospital Basel, Basel, Switzerland. 56: Johns Hopkins University School of Medicine, Department of Pathology, Division of Neuropathology, Baltimore, MD, USA 57: Section of Neuropathology, Department of Pathology and Laboratory Medicine, and Department of Neurology, Brain Research Institute, University of California, Los Angeles (UCLA) Medical Center and David Geffen School of Medicine, Los Angeles, California, USA 58: Laboratory of Neuropathology, Department of Pathology and Neuropathology, Neuromed Campus, Kepler University Hospital, Medical School, Johannes Kepler University, Linz, Austria 59: Center for Cognitive Neurology, Departments of Neurology, Pathology and Psychiatry, New York University School of Medicine, ERSP, 450 East 29th Street, NY, NY, USA 10016 60: Centre for Cancer Therapeutics, Ottawa Hospital Research Institute, Department of Pathology and Laboratory Medicine, University of Ottawa 61: Department of Neurology & Neurobiology of Aging, Kanazawa University Graduate School of Medical Sciences, Kanazawa, Japan

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Word count abstract: 296

Word count text: 4796

Number of Figures: 4

Number of tables: 3

Corresponding authors: Gabor G. Kovacs MD PhD

Institute of Neurology, AKH 4J, Währinger Gürtel 18-20, A-1097 Vienna, Austria

Phone: +43-1-40400-55070; Fax +43-1-40400-55110;

Email: gabor.kovacs@meduniwien.ac.at

and

Dennis W. Dickson MD

Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL 32224,

USA

Email: dickson.dennis@mayo.edu

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Abstract

Pathological accumulation of abnormally phosphorylated tau protein in astrocytes is a

frequent, but poorly characterized feature of the aging brain. Its etiology is uncertain, but its

presence is sufficiently ubiquitous to merit further characterization and classification, which

may stimulate clinicopathological studies and research into its pathobiology. This paper aims

to harmonize evaluation and nomenclature of aging-related tau astrogliopathy (ARTAG), a

term that refers to a morphological spectrum of astroglial pathology detected by tau

immunohistochemistry, especially with phosphorylation-dependent and 4R isoform specific

antibodies. ARTAG occurs mainly, but not exclusively, in individuals over 60 years of age.

Tau-immunoreactive astrocytes in ARTAG include thorn-shaped astrocytes at the glia

limitans and in white matter, as well as solitary or clustered astrocytes with perinuclear

cytoplasmic tau immunoreactivity that extends into the astroglial processes as fine fibrillar or

granular immunopositivity, typically in gray matter. Various forms of ARTAG may coexist in

the same brain and might reflect different pathogenic processes. Based on morphology and

anatomical distribution, ARTAG can be distinguished from primary tauopathies but may be

concurrent with primary tauopathies or other disorders. We recommend four steps for

evaluation of ARTAG: 1) identification of five types based on the location of either

morphologies of tau astrogliopathy: subpial, subependymal, perivascular, white matter, gray

matter; 2) documentation of the regional involvement: medial temporal lobe, lobar (frontal,

parietal, occipital, lateral temporal), subcortical, brainstem; 3) documentation of the severity

of tau astrogliopathy; and 4) description of subregional involvement. Some types of ARTAG

may underlie neurological symptoms; however, the clinical significance of ARTAG is

currently uncertain and awaits further studies. The goal of this proposal is to raise awareness

of astroglial tau pathology in the aged brain, facilitating communication among

neuropathologists and researchers, and informing interpretation of clinical biomarkers and

imaging studies that focus on tau-related indicators.

Key words: aging; ARTAG; tau astrogliopathy; tau;

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Introduction

Tau is a microtubule-associated protein that binds to tubulin and promotes its polymerization

and stabilization into microtubules. Tau isoforms, ranging from 352 to 441 amino acids, are

generated by the alternative splicing of exons 2, 3, and 10 of the MAPT gene. The six

isoforms differ from each other by the presence or absence of 29- or 58-amino acid inserts in

the N-terminus domain and by the presence of either three (3R tau isoforms) or four (4R tau

isoforms) tandem repeat sequences of 31 or 32 amino acids [24]. Mutations in the tau gene

(MAPT) can cause hereditary frontotemporal dementia and associate with frontotemporal

lobar degeneration (FTLD) [23, 26, 51, 63]. Following the description of a disorder in one

family named ‘multiple system tauopathy with presenile dementia’ [62], the term tauopathy

was introduced to refer to disorders in which tau protein deposition is the predominant feature

[23]. Tauopathies are characterized by the accumulation of abnormal and hyper-

phosphorylated tau protein in the brain and are also classified as primary or secondary [32,

37]. Tau pathology is characterized as 3R or 4R predominant or mixed 3R+4R type [12, 30,

32]. Primary tauopathies are grouped also as FTLD-tau and comprise Pick disease (PiD),

progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), argyrophilic grain

disease (AGD), neurofibrillary tangle (NFT) predominant senile dementia (NFT-dementia or

“tangle-only” dementia; now included in the category of PART, see below), and globular glial

tauopathy (GGT) [32, 47]. In addition, many other diseases or conditions with diverse

etiology, including Alzheimer disease (AD), may be associated with tau pathology [32]. The

recently introduced term 'primary age-related tauopathy' (PART) encompasses neuronal

changes previously considered as “normal aging” as well as NFT-dementia [14]. PART is

distinguished from AD, largely by the absence or scarcity of amyloid (Aβ) plaques [14]. In

aged individuals sex-dependent tau pathology, developing independently from AD has been

also described in the hypothalamus [16, 54, 56]. Furthermore, chronic traumatic

encephalopathy (CTE) is associated with a distinctive pattern of progressive neuronal and

glial tau pathology [40-42].

The introduction of the Gallyas silver stain and particularly diagnostic tau

immunohistochemistry led to the identification of astroglial tau pathology in the aging brain

in people with or without AD-related changes, cognitive decline or movement disorders [5, 7,

11, 21, 25, 31, 34-36, 38, 46, 57]. There have been attempts to classify these tau

astrogliopathies [34], but there is lack of consensus as to how best to describe and categorize

them. We recommend the term aging-related tau astrogliopathy (ARTAG) to describe the

7

spectrum of otherwise unclassified tau immunoreactivity in astrocytes (i.e., distinct from

tufted astrocytes, astrocytic plaques, ramified astrocytes, or globular astroglial inclusions)

mostly in aged individuals detected by tau immunohistochemistry using phosphorylation-

specific, conformation-specific, or isoform-specific (4R) anti-tau antibodies. Both ARTAG

and PART affect predominantly the elderly, but PART is characterized by neurofibrillary

degeneration that is largely restricted to the medial temporal lobe (MTL), basal forebrain,

brainstem, and olfactory bulb and cortex [14]. PART thus describes neuronal tau pathology,

while ARTAG focuses on astrocytic tau pathology. Whether PART and ARTAG belong to

separate or shared pathogenic processes is unknown.

We propose a four-step approach for the morphological classification of ARTAG. We

anticipate that harmonizing the nomenclature and improving consistency in documentation of

ARTAG is a necessary first step for defining diagnostic guidelines that will result in progress

in clinicopathological correlation and investigation of the pathogenesis of ARTAG.

Morphology of tau-immunoreactive astrocytes in primary tauopathies and CTE

The defining lesions of tauopathies are intracellular aggregates of abnormal conformers of

tau, consistently detectable by immunohistochemistry for phosphoepitopes (e.g., PHF1, CP13,

and AT8), as well as epitopes to conformational epitopes (e.g., Alz50 and MC1) and tau

isoform specific epitopes (e.g., 4R tau isoforms) [32]. Regardless of the tauopathy, astroglial

tau inclusions are mostly 4R tau-immunopositive, although ramified astrocytes in PiD as well

as occasional protoplasmic astrocytes in PSP may show 3R-tau immunoreactivity [21]. Tufted

astrocytes are characteristic of PSP, and astrocytic plaques are signature lesions of CBD,

while so-called ramified astrocytes have been described in PiD [15, 32]. In addition,

astroglial, argyrophilic, and intracytoplasmic flame or thorn-shaped inclusions were described

by Nishimura et al. in PSP [49]. Phosphorylation-dependent anti-tau antibodies are highly

sensitive and label lesions that are not consistently detectable by silver impregnation methods,

but may show variable ubiquitin or p62/sequestosome immunopositivity, such as the globular

astroglial inclusions (GAI) of GGT [1], or the fine granular tau immunopositivity (some with

‘bush-like’ appearance) of the astrocytes of AGD [11]. Some of the variation in the

morphology of the immunolabeled structures was interpreted as representing stages of a

process of aggregation and fibrillation, analogous to progression from pretangles to

neurofibrillary tangles in AD [6, 11]. The concept of early-stage tau accumulation in

astrocytes has also been discussed in relation to the changes in the basal ganglia in PSP [53].

8

Finally, subpial and subependymal clusters of astrocytic tangles have been described in CTE

[41].

Overview of astrocytic tau pathologies in the aging brain

Both neuronal and glial tau pathology increases in frequency with age. The most frequent

neuronal tau inclusions are neurofibrillary tangles, threads, and argyrophilic grains. Neuronal

and glial inclusions resembling PSP pathology can be seen in the elderly, even without

clinical evidence of PSP [17, 18, 34], but these lack the typical multisystem degeneration seen

with PSP. Furthermore, tuft-shaped astrocytes have been described in a subgroup of elderly

individuals, especially in association with Lewy body pathology, in a distribution resembling

that of PSP [25]. Nevertheless, converging data emphasize the presence of a tau

astrogliopathy that differs from tufted astrocytes or astrocytic plaques as a common finding in

the elderly. Despite its high prevalence, there is a lack of consensus on whether these

astroglial tau pathologies in the elderly are clinically relevant, even as a concomitant

pathology that might lower an individual’s threshold for the development of clinical

symptoms. Research in this field has been hampered by the variation in staining methods, tau

antibodies, and the inconsistent nomenclature for astroglial tau pathologies. Importantly,

hypertrophic astrocytes, as revealed by hematoxylin and eosin staining and

immunohistochemistry for glial fibrillary acidic protein and excitatory amino acid transporter

2 (EAAT2), are common in the elderly, and presumably represent a reaction to multiple types

of injury. The location of such reactive astrocytes varies considerably among individuals [8,

59]. Colocalization studies have indicated that glial fibrillary acidic protein-immunoreactive

reactive astrocytes are not necessarily those that are also immunoreactive for tau pathology

[19].

Ikeda and colleagues were the first to describe tau-positive thorn-shaped astrocytes (TSA),

which were similar in morphology to tau-positive astrocytes described by Nishimura et al. in

PSP [49]) in the subpial or subependymal regions of the gray and white matter and frequently

in the depths of gyri, as well as in the basal forebrain and brainstem, in aged individuals [27-

29]. TSA may occur in multiple conditions [13]. In comparison to the tufted astrocytes of PSP,

TSA showed more voluminous perinuclear cytoplasm and their processes are often thicker

and shorter [27]. TSA were only occasionally found in the deep cortical layers. The authors

noted that anti-ubiquitin antibodies do not label TSA. They interpreted TSA as a non-specific

finding and found no relationship between the number of TSA and the severity of

9

neurofibrillary changes. Argyrophilic, tau-positive subpial and perivascular structures were

also described as common TSA [27-29].

Schultz et al. reported a high prevalence of TSA in the aged human MTL, particularly the

anterior MTL, at the level of the amygdala [57]. TSA were absent in individuals under 60

years, but affected almost half of brains from those over 75 years [57]. Indeed, another study

also failed to find this type of tau astrogliopathy in younger individuals [33]. Schultz et al.

[57] commented that tau immunopositivity was not confined to the thorn-shaped proximal

processes of astroglia, but also presented as thread-like processes in the neuropil. They found

that immunolabeling with AT8 was the most sensitive for demonstrating the TSA, while

silver staining was less consistent [57]. They also speculated that the preferential subpial and

perivascular location could be a result of exposure to CSF or to extravasated plasma proteins

due to defects in blood-brain barrier permeability commonly seen in aging and

neurodegeneration [36, 57]. Interestingly, a similar distribution of TSA was reported in aged

baboons [55]. A study by the MRC-CFAS group confirmed the findings of Schultz et al. and

added that the TSA could be less commonly observed in the vicinity of neuronal cell bodies in

gray matter areas such as amygdala and dentate gyrus [36]. Also, this study documented the

4R tau nature of TSA [36]. Variable staining for Gallyas and p62 suggest that some of these

astrocytes accumulate tau in a fibrillar state [34-36, 38, 46]. Uchikado et al. also reported that

the frequency of TSA increased with age and was independent of AGD [66]. All studies agree

that the burden of TSA is independent of AD pathology, AGD, coiled bodies, dementia status

at death, or presence of the APOE ε4 allele [27, 28, 36, 57, 66]. These studies, however, were

limited to evaluation of the MTL and did not take account of cortical and subcortical tau

astrogliopathy. Few studies reported tau immunopositivity in glial cells in AD cases with

prolonged duration of the disease [5, 48, 68]. Finally, a report on NFT-dementia mentioned

the presence of astrocytic tau pathology in white matter and cortex [31].

The possibility that TSA may have clinical significance was first raised by Munoz and

colleagues [46]. They used the term ”argyrophilic thorny astrocyte clusters (ATACs)” and

observed them in the frontal, temporal, and parietal cortices and in subcortical white matter in

a cohort of patients with nonfluent variant of primary progressive aphasia associated with AD

pathology [46]. Subsequent reports also linked TSA to symptomatology, although not all

found an association of ATACs with focal neurological syndromes [9, 43]. Munoz and

colleagues noted ATACs in the white matter without discernible changes in sections stained

10

for myelin. ATACs did not show a consistent topographic relationship to amyloid deposits,

NFTs, or reactive astrogliosis [46]. Interestingly, focal glial tauopathy, interpreted as PSP-

type, associated with progressive aphasia was reported by Wakabayashi et al. [67]. These

observations suggested that TSA-like astrocytes might be detected not only in a

subependymal or subpial location.

A peculiar constellation of tau pathology was documented in a subset of patients with

dementia [35]. The most characteristic feature was a tau astrogliopathy, which was described

as diffuse granular tau immunoreactivity in astrocytic processes [35]. The study emphasized

additional neuronal pathologies, including threads and diffuse neuronal cytoplasmic tau

immunoreactivity (pretangle-like). A further study distinguished four different patterns based

on the anatomical distribution of the tau astrogliopathy and its combination with neuronal tau

pathology, characterized mostly by pretangles and scattered threads [34]: 1) medial temporal

lobe type (Group I); 2) amygdala type (Group II); 3) limbic-basal ganglia-nigral type with

neuronal tauopathy (Group III); and 4) hippocampus-dentate gyrus-amygdala type with

neuronal tauopathy (Group IV). Some of these might represent stages of the same process

whereas other might be different. Nevertheless, evaluation of tau astrogliopathy in several

anatomical regions indicated that in some cases astroglial tau pathology in the elderly extends

beyond the MTL to involve the frontal and parietal cortices, striatum, substantia nigra, and

medulla [34]. The morphology of tau astrogliopathy in these studies, was reminiscent of that

reported by Munoz et al. as ATACs [46], although extension of the immunoreactivity into the

astrocytic processes was emphasized [34, 35]. Distinct accumulation of TSA in the dentate

gyrus of the hippocampus was also recognized [34, 36]. Mathematical modeling of

hippocampal tau immunolabeling patterns suggested that some forms of tau astrogliopathy in

the elderly involve hippocampal subregions in a different pattern from that of primary

tauopathies [44]. Ferrer et al. [21, 38] showed that the biochemical signature of astroglial tau

pathology in the elderly in both white and gray matter differed from that of other astrocytic

tau pathologies in primary tauopathies. Specifically, astroglial tau pathologies in the white

matter and gray matter in aging brains were not consistently detectable using phospho-

specific anti-tau antibody Ser262 or conformational tau modifications at amino acids 312 to

322 (MC1), or tau truncated at aspartic acid 421 (tau-C3) [21, 38].

In addition, isolated tufted astrocytes were reported in the occipitotemporal gyrus in an

elderly, population-representative cohort [36], and a tauopathy with tufted, thorny, fibrous,

and protoplasmic forms of astrocytic pathology was described by Beach et al. [7], in a series

11

of cases with hippocampal sclerosis and also in a community-based study [34]. Sakai et al.

[52] reported prominent subcortical white matter astrocytic tau pathology in brains from two

elderly patients in whom CBD was considered. In a study on cervical spondylotic

myelopathy, AT8 immunohistochemistry revealed tau-positive, neuropil threads, astrocytic

foot-like perivascular or subpial structures, and glial cells with short and thick processes,

which the authors termed TSA [58]. Interestingly, prominent tau astrogliopathy may be seen

in familial disorders without MAPT mutation [20].

In summary, tau-immunoreactive astrogliopathy in the elderly represent a spectrum of

morphological abnormalities including those originally described as TSA (plump, perinuclear

cytoplasmic immunoreactivity) and additional fine granular tau immunoreactivity extending

into the astrocytic processes in the gray matter. These two morphologies can be present in the

same brain. TSA may be seen in subpial, subependymal, or perivascular areas, as well as in

the white and gray matter, while the fine granular immunoreactivity is seen in the gray matter.

Most likely, the different tau-immunoreactive astroglial morphologies in different locations in

the aging brain, with or without clinical correlation, reflect different pathogenetic events. We

propose the umbrella term ARTAG to encompass all of these, with or without accompanying

morphological features of other neurodegenerative disorders, including PSP, CBD, PiD,

GGT, PART, AD, AGD, and Lewy body pathology. Some clinicopathological studies suggest

that ARTAG may present clinically with focal symptoms like aphasia when circumscribed to

a smaller number of regions [46], whereas, in cases with widespread pathology dementia with

or without parkinsonism might be the clinical presentation [34, 35]; on the other hand, studies

focusing only on the MTL have found no relationship between ARTAG and cognitive

impairment or dementia [36].

Differential diagnosis

We provide the following operational criteria for the six well-defined tau immunoreactive

astrocytic cytopathologies seen in primary tauopathies and ageing brain as follows (see

comparison in Table 1 and Fig. 1):

1) Tufted astrocytes: star-like tufts of tau-positive radiating fibers. The dense tau

immunoreactive tufts are detected in the proximal part of the astrocytic processes, often

usually in a symmetrical fashion. They are localized to the gray matter (mostly basal ganglia

and neocortex).

2) Astrocytic plaques: focal and densely tau-immunoreactive stubby dilatations of distal

12

processes of astrocytes giving a senile-plaque-like appearance without amyloid core. They are

localized to the gray matter (mostly basal ganglia and neocortex).

3) Ramified astrocytes: tau immunoreactivity occupying mostly the perikarya and ramifying

into the cell processes usually localized to one side of the cell giving the appearance of

eccentric nuclei of the astrocyte. They are localized to the gray matter and to the white matter

in neocortices with severe neuronal loss.

4) Globular astroglial inclusions: tau immunoreactive distinct globules (up to the size of the

astroglial nucleus; 1-5 µm) and dots (1-2 µm) in the perikarya and proximal parts of

astrocytic processes, found in gray matter.

5) Thorn-shaped astrocytes (TSA): tau immunoreactivity is localized in astrocytic perikarya

with extension into the proximal parts of the astrocytic processes, with inclusions also in the

astrocytic endfeet at the glia limitans around blood vessels and at the pial surface. The

processes are thick and short and reminiscent of thorns. They are preferentially found at

subpial and perivascular locations, as well as in the white matter and less often as clusters in

the gray matter.

6) Granular or fuzzy tau immunoreactivity in processes of astrocytes (GFA): fine granular

immunoreactivity of branching processes with a few dilations of gray matter astroyctes. The

perinuclear soma is densely immunoreactive in most of these astrocytes.

The two major cytomorphologies of ARTAG (i.e., TSA and GFA) may accompany

tauopathies or other neurodegenerative disorders, but ARTAG should be distinguished from

the more specific astrocytic lesions that are characteristic of primary tauopathies. To

understand the frequency and relevance of ARTAG we recommend documenting ARTAG as

an additional feature in primary tauopathies. It must be noted, that the astroglial tau

immunoreactivity described by Botez et al. in the amygdala of AGD [11] fits best with the

GFA now included as a form of ARTAG. Indeed, astrocytic tau pathology is variably seen in

AGD [22]. Therefore, it is helpful to comment whether in a case of AGD additional ARTAG

is present. Furthermore, there are other tau-related disorders with astrocytic tau pathology.

For instance, astrocytic tau pathology is also a component of CTE [40-42]. CTE is associated

with a history of repetitive concussive or subconcussive brain trauma and is characterized by

widespread accumulation of hyperphosphorylated tau in NFTs and astrocytes, which have

similarity to TSA seen in ARTAG [41]. ARTAG has features that overlap those of CTE,

including the accentuation of tau pathology around small cerebral vessels and in subpial and

periventricular areas. On the other hand, tau pathology, including neuronal and astroglial, in

13

CTE is more abundant in the depths of the convexity cerebral sulci, especially in early stages

[41], an aspect that has not been reported in tau astrogliopathy in the aging brain [29, 34-36,

46, 57]. It is possible that CTE pathology has been considered to be age-related

astrogliopathy, especially for lesions in the MTL, which can be severely affected in more

advanced stages of CTE [42]. The characteristic patchy lesions at depths of cerebral sulci

were not recognized as a specific morphological feature of CTE in earlier studies. Finally,

tufted astrocytes in PSP, astrocytic plaques in CBD, globular astrocytic inclusions in GGT,

and ramified astrocytes in PiD are distinct from tau-immunoreactive astrocytes in the gray

matter in ARTAG (see Table 1).

These observations raise the possibility that ARTAG affects distinct astrocytic populations to

those in established primary tauopathies. Ikeda and colleagues noted that the distribution of

TSA was coexistent with prominent subpial and subependymal gliosis [29]. Corpora

amylacea, which are heavily invested by reactive astrocytes, also share this distribution.

Importantly, these astroglial populations of the “glia limitans” share common features with

fibrous astrocytes, which predominate in the white matter and subpial zone [8] and with a

subset of astrocytes in the gray matter [61], where ARTAG can be also observed. In contrast,

astrocytic tau pathologies in CBD or PSP involve protoplasmic astrocytes and are

independent of reactive astrogliosis [19, 65]. A few studies report association of glial

fibrillary acidic protein and AT8 immunoreactivity in subpial but also in gray matter

localization of tau astrogliopathy in elderly brains [35, 36]. Protoplasmic and fibrous

astrocytes differ substantially in their glutamate uptake capabilities and capacity and have

very different degrees of coupling, which are important with regard to their respective

calcium wave signals, resting membrane potentials, potassium buffering, glutamate

metabolism, exchange of second messengers, metabolites, and other signaling intermediates

between cells [50]. In addition to these differences, reaction of astrocytes varies considerably

between distinct diseases of the nervous system [60]. It is these differences that may be of

pathogenetic relevance to the morphologic diversity of astrogliopathy in ARTAG.

Evaluation of ARTAG

Inconsistency in assessing, describing and documenting ARTAG has impeded research and

limited our understanding of the significance of this pathology. It is not clear whether the

different patterns of anatomical involvement represent a continuum or distinct abnormalities

with different causes. Most previous studies have focused on the MTL, but more widespread

14

involvement is possible [34, 35]. The relative frequency of ARTAG limited to MTL as

opposed to more widespread tau astrogliopathy remains unclear. Potential etiologies are not

known, although defective function of the blood-brain barrier [57], metabolic encephalopathy,

neurodegenerative pathologies, hypoperfusion associated with aging, AD, or vascular

dementia [39, 64], and even repeated minor trauma with possible genetic risk factors may

play a role. Clinical, imaging and neuropathological data related to these aspects need to be

documented precisely to allow a better understanding of the pathogenesis of ARTAG [47]. A

method is needed to describe morphologies that can be widely accepted and reproducible. As

silver impregnation methods are difficult to standardize and immunohistochemistry for

ubiquitin and p62 does not demonstrate all forms of tau cytopathology, optimal

characterization of ARTAG requires the use of immunohistochemistry for phosphorylated

tau. The most widely used phosphorylation-dependent anti-tau antibodies that have allowed

characterization of ARTAG to date include: AT8 (pSer202/Thr205; available from different

commercial sources), CP13 (Ser202; Peter Davies, Litwin-Zucker Research Center for The

Study of Alzheimer’s Disease and Memory, Manhasset, NY, USA) and PHF-1 or AD2

(Ser396/Ser404; Peter Davies, NY, USA for PHF-1 or commercial sources for AD2s). Other

antibodies that may prove useful in the characterization of ARTAG include those specific for

tau phosphorylated at Thr181, Ser202, Ser214, Ser396, Ser422, N-terminus region epitope-

specific, 4R tau isoform-specific, and some conformation-dependent antibodies such as Alz50

(but not MC-1) [21, 35, 38].

Recommendations for sampling and staining are as follows:

- Preliminary screening for ARTAG should include tau-immunohistochemistry (antibodies

AT8, CP13, AD2 or PHF-1 are recommended) on two sections representative of the MTL

(i.e., amygdala and hippocampus at the level of the lateral geniculate body). These regions

are vulnerable to TSA and GFA.

- If tau astrogliopathy is noted in the screening section, a systematic characterization of

ARTAG will require analysis involving additional areas of the frontal, parietal, lateral

temporal, and occipital cortices, as well as anterior and posterior portion of the basal

ganglia, thalamus, midbrain at the level of substantia nigra, pons at the level of locus

coeruleus, and medulla oblongata.

- In cases where focal cortical symptoms are reported, further cortical areas corresponding to

the clinical symptoms or signal alterations detected in MRI should also be evaluated.

15

ARTAG should be considered when detecting either or both of the two cytomorphologies: TSA

or GFA. As such, we propose the following four-step characterization TReSS algorithm (Table

2):

Type? Regional involvement? Severity? Subegional involvement?

• First: Identify the morphologic and distribution types of ARTAG based on parenchymal

localization of TSA and GFA (note that combination of these types is generally the rule):

1) Laminar subpial TSA (Fig. 2a): plump perinuclear cytoplasmic tau

immunoreactivity in astrocytes in subpial locations. It is important to note whether this

is more pronounced in the sulcal depths in the convexity cerebral cortices, as in CTE.

2) Subependymal TSA: plump perinuclear cytoplasmic tau immunoreactivity in

astrocytes in subpial or subependymal locations (Fig. 2b).

3) Perivascular TSA: plump cytoplasmic immunoreactivity with tau-immunoreactive

astrocytic processes around vessels (Fig. 2c) in the gray or white matter.

4) White matter TSA: astrocytes in the subcortical white matter that show plump

cytoplasmic immunoreactivity (Fig. 2d). Note that in the white matter these usually

form small clusters (>3 astrocytes) and that it may extend into the adjacent gray matter

as described for ATAC [46].

5) Gray matter GFA: solitary (one or two/20x field) (Fig. 2e, f) or clustered (Fig. 2g,

h) GFA (three or more/20x field) with fine granular immunopositivity of the

cytoplasmic processes (GFA), with plump perinuclear cytoplasmic tau

immunoreactivity. Less frequently, TSA can be also seen in the gray matter.

It must be noted that tau immunohistochemistry occasionally decorates astrocytes at

the border of chronic vascular lesions in young and aged individuals. Therefore, this

lesion-asssociated tau astrogliopathy is important to document, but is not considered

an aging related astrogliopathy.

• Second: Identify involvement of gross anatomical regions:

A) MTL

B) Lobar

C) Subcortical

D) Brainstem

16

Although the most frequently involved region is the MTL, involvement of further

regions should be recognized. Moreover, MTL is important for comparison with

neuroimaging data on MTL atrophy.

• Third: Document the severity of ARTAG pathology in the region or subregion (see step

four) examined. ARTAG may appear in focal clusters or in a widespread distribution. We

propose documentation as to whether ARTAG involves only 1) occasional or 2)

numerous astrocytes. If the latter, the focal clusters or widespread distribution should be

noted. Semiquantitative scoring for ARTAG will need to be refined.

• Fourth: Map subregional involvement to promote future exploration and scientific

discovery related to ARTAG. These are the subdivisions within the gross anatomical

regions of the second step and include the following (Table 3; examples are shown in Fig.

3):

- amygdala and hippocampus, inferior temporal gyrus for MTL

- frontal, parietal, occipital, lateral temporal (middle and superior gyrus) for lobar

- caudate nucleus, putamen, nucleus accumbens, globus pallidus, thalamus, basal

forebrain for subcortical

- mesencephalon, pons, medulla oblongata for brainstem

It should be noted whether ARTAG is associated with features of a particular neurodegenerative

disorder, or with other disease (cerebrovascular, inflammatory, metabolic, etc.) followed by the

description of the type and major regional involvement.

Some examples for the diagnostic reporting are provided as follows:

1) Examples for pure types:

a. ARTAG subpial type;

Region: MTL;

Subregion: hippocampus, inferior temporal cortex;

Extent: numerous astrocytes and widespread distribution

b. ARTAG subependymal type;

Region: Subcortical;

Subregion: lateral ventricle;

Extent: occasional

17

2) Examples for combinations:

a. ARTAG gray matter type;

Region: MTL and subcortical;

Subregion: inferior temporal cortex and nucleus accumbens;

Extent: numerous astrocytes in focal clusters

plus

ARTAG white matter type;

Region: MTL;

Subregion: hippocampus, periamygdala white matter, and temporal;

Extent: numerous astrocytes with widespread distribution;

b. ARTAG perivascular type;

Region: lobar and subcortical;

Subregion: frontal cortex and striatum;

Extent: occasional

plus

ARTAG white matter type;

Region: MTL and lobar;

Subregion: lateral temporal, frontal, and parietal lobes;

Extent: numerous astrocytes in focal clusters.

For example, the cases described by Munoz et al. [46] would be summarized in the diagnostic

report as: ARTAG gray and white matter type; region: MTL and lobar; extent: numerous in

focal clusters. For research purposes, the subregional involvement should be added as: gyrus

ambiens, parahippocampal gyrus, fusiform gyrus, inferior, middle, and superior temporal gyri,

frontal dorsolateral and orbitofrontal cortices, cingulate gyrus, and inferior parietal lobe. The

cases described by Kovacs et al. [35] could be summarized as ARTAG gray matter type;

region: MTL, lobar, subcortical, and brainstem; extent: numerous in focal clusters; plus white

matter type; region: MTL; extent: numerous and widespread; with further details on the

subregional involvement. The cases discussed by Santpere and Ferrer [53] as early PSP-like

astrocytic changes also represent ARTAG with additional features of concomitant PSP-type

pathology (i.e. cases 4 and 5).

18

Summary

ARTAG describes a spectrum of astroglial tau pathologies detected mainly in the elderly

represented by TSA and GFA, which are distinct from astroglial lesions of primary

tauopathies (i.e. tufted astrocytes, astrocytic plaques, ramified astrocytes, or globular

astroglial inclusions). The frequency of ARTAG varies depending on the type: subpial,

subependymal, and perivascular types are more frequent, while gray matter and cerebral white

matter types might be less common. The etiology of different types might be different;

however, all appear mostly associated with aging. Although documented in several

publications, there is a lack of consensus on how ARTAG should be recorded and interpreted.

Here we propose steps for a systematic characterization with the expectation that this will

improve communication about and understanding of this condition, including its relation to

other brain pathologies and clinical symptoms. This approach has the potential to help in

several respects:

1) It will facilitate communication between neuropathologists and

researchers. Revisiting and standardizing the terminology should help to move the

field forward. It will also increase awareness of this pathology, which is under-

recognized and under-studied.

2) A better differentiation of ARTAG types may help with assessing their relationship

to other tauopathies. This may be particularly important in the context of CTE-related

tau pathologies. Furthermore, this may help better understanding of differences in the

pathogenesis of ARTAG types.

3) A regular system for typing and grading of ARTAG should facilitate comparisons

between different centers, and the pooling of information in harmonized

clinicopathological studies. These will potentially pave the way towards mechanistic

insights and genetic studies into their pathogenesis.

4) Understanding the nature of ARTAG may help in the interpretation of clinical

biomarker and imaging studies.

Development of such a common concept (Fig. 4) and nomenclature that allows comparisons

across studies and aggregation of data for large scale multi-institutional analyses is imperative

in order to understand the phenomena and clinical implications of ARTAG. Future studies

should also aim to re-evaluate these observations to validate this approach and to develop a

concise classification of ARTAG for diagnostic neuropathology. To reach this goal,

19

paradigms will need to be designed for ARTAG along the lines used to standardize evaluation

and diagnostic criteria for tau, amyloid and α-synuclein and other major pathologies [2-4, 10,

45]. Subsequently, it will be possible to evaluate inter-rater reliability of the proposed

evaluation and eventually to merge clinical and pathologic data from multiple centers to

determine practical significance of ARTAG. At this preliminary stage, however, our

recommendation is limited to an evaluation strategy focusing primarily on common

nomenclature and classification of aging related astrogliopathy.

20

Acknowledgements

We are extremely grateful to the patients, clinicians, and fellow researchers that made this

effort possible. We also acknowledge the following funding sources: FP7 EU Project

Develage No. 278486 (GGK); Grant "NIH P30 AG10133 (BG); NIA grants P50 AG05681,

P01 AG03991 (NJC), NIH R01 AG040311, institutional grants NIH P01 AG019724-03 and

P50 AG023501, and the tau consortium (LTG); the Nelson Family Foundation and NIH

grants AG010124, AG017586 (MEM); NIH grant P50 AG005138 (PRH); Alzheimer's

Research UK (ARUK), Alzheimer's Society, National Institute for Health Research (NIHR),

and UK Medical Research Council (MRC; G0400074)(JA); GMH is a National Health and

Medical Research Council of Australia Senior Principal Research Fellow (#630434); grant

IGA NT12094-5 from Grant Agency of Ministry of Health of Czech Republic (RM); NIH

grant # AG028383 (PN); UK Medical Research Council (MRC; MR/L016400/1) (CS); NIA

P50 AG005133 (J.K); National Institute of Neurological Disorders and Stroke

(1U01NS086659-01), Department of Veterans Affairs, ), the National Institute of Aging

Boston University Alzheimer’s Disease Center (P30AG13846; supplement 0572063345–5)

(ACM); UK Medical Research Council (MC-PC-13044) (JWI and CS); National Brain

Research Program, Hungary (KTIA_13_NAP-A-II/7) and Grant-in-Aid (KAKEN

26250017)(both for TH); NIH grant P30AG12300 (KH, CLW); Ministerio de Ciencia e

Innovación, Instituto de Salud Carlos III – Fondos FEDER, a way to build Europe FIS grants

PI14/00757 and PI14/00328 (IF); DFG Grant (SFB 1134/A03)(CS); Johns Hopkins

Alzheimer's Disease Research Center NIH grant #P50AG05146 (JCT); Alzheimer's Disease

Core Center grant P30AG008051-26 (TW); Grant AG13854 (EHB); JSPS KAKENHI Grant

Number� 26430060 (MT); Italian Ministry of Health (GG and FT); National Institute of

Health grants P50 AG05136 and P50 NS062684 (TJM). The help of Brain Banks in collecting

tissue is also highly acknowledged: Vienna KIN-Neurobiobank and VITA–study (GGK); GIE

NeuroCEB (funded by the patients associations France Alzheimer, France Parkinson,

Fondacion ARSEP and CSC)(CD); Sydney Brain Bank (funded by Neuroscience Research

Australia and the University of New South Wales)(GMH); the Sheffield and Cambridge Brain

Banks (CFAS)(PI, SW); Parkinson's UK Tissue Bank at Imperial College, funded by

Parkinson's UK, a charity registered in England and Wales (948776) and Scotland

(SC037554)(SG); The Edinburgh Brain Bank is supported by the UK Medical Research

Council (MR/L016400/1)(CS, JWI).

21

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25

Table 1. Operational criteria for the six well-defined tau-immunoreactive astrocytic cytopathologies seen in primary tauopathies and aging-related tau astrogliopathy (ARTAG))(see also Fig.1). GM: gray matter; WM: white matter; SP: subpial; SE: subependymal; PV: perivascular. In the column "Reported” those diseases are mentioned, where there are literature reports of the specific astrocytic tau pathologies. PSP: progressive supranuclear palsy; CBD: corticobasal degeneration; CTE: Chronic Traumatic Encephalopathy; PiD: Pick disease; GGT: globular glial tauopathy; AGD: argyrophilic grain disease. For Gallyas silver staining: + indicates consistently detectable; -/+ indicates variably detectable (for GFA the soma may be variably positive but the processes not); - indicates usually not detectable.

Characteristic immunoreactivity Astrocytic processes Name Soma Proximal Distal Gallyas Reported Location a. Primary tauopathy-related astroglial tau pathology

Tufted astrocytes

relatively spared

dense tufts: usually

symmetric no + PSP GM

Astrocytic plaques spared no stubby

dilatations + CBD GM

Globular astroglial inclusions

dense: filled with globules

1-5 µm globules

1-2 µm globules - GGT GM

Ramified astrocytes

dense: localized to

one side

dense ramifications:

usually asymmetric

no + PiD GM,

WM/GM junction

b. ARTAG-related astroglial tau pathology

Thorn-shaped astrocytes (TSA)

dense: thorn or flame shaped

short and thick no -/+

with or without

PSP, AGD, AD, CTE,

other

GM, WM, SP, SE, PV

Granular or fuzzy astrocytes (GFA)

dense: perinuclear

accumulation

fine or fuzzy granular

fine granular -/+

with or without AGD

GM

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Table 2. Evaluation of aging-related tau astrogliopathy (ARTAG). TSA: thorn-shaped astrocyte; GFA: granular /fuzzy astrocyte; AP: astrocytic plaque; TA: tufted astrocyte; RA: ramified astrocyte; GAI: globular astroglial inclusions.

Requires: Presence of thorn-shaped astrocytes (TSA) and/or solitary or clustered astrocytes with plump cytoplasmic tau immunoreactivity that extend into the astroglial processes as fine granular immunopositivity (GFA) distinguishable from AP, TA, RA, and GAI.

Four-step characterization of ARTAG:

Step 1: Distinguish types according to the location:

Subpial

Subependymal

Gray matter

White matter

Perivascular

Step 2: Describe major anatomical distribution

Medial temporal lobe

Lobar

Subcortical

Brainstem

Step 3: Document the severity of ARTAG

Occasional

Numerous

- Focal

- Widespread

Step 4: Map subregional involvement and extent (see Table 3)

Ancillary studies: - Description of additional tau pathologies in specific anatomical regions:

• Neurofibrillary degeneration

• Neuropil threads

• Diffuse cytoplasmic neuronal tau immunoreactivity (“pretangles“) • Argyrophilic grains

• Dystrophic neurites around or within amyloid plaques

• Oligodendroglial tau immunoreactivity - Characterization of tau phosphorylation, conformation, truncation, nitration, ubiquitination, immunohistochemistry for 4R and 3R tau; ultrastructural study; genetic studies (MAPT and other genes associated with neurodegeneration)

- Description of concomitant neurodegenerative and non-neurodegenerative pathologies

- Description of relation to lesions (”perilesional” astrocytic tau immunoreactivity), to corpora amylacea, and Rosenthal fibers

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Table 3. Description of aging-related tau astrogliopathy (ARTAG) based on the type and distribution of astrocytic tau pathology. MTL: medial temporal lobe; Gy: gyrus; GP: globus pallidus; Caud/Put: Caudate and Putamen; Dent Gyr: dentate gyrus; Medulla obl.: medulla oblongata; Aq: Aqueduct; LV: lateral ventricle; 3V: 3rd Ventricle. *: in the case of focal cortical symptoms the anatomical area with clinical relevance should be noted additionally. Combinations of subtypes should be expected and described.

Diagnostic screening

Clinicopathological correlation and studies on pathogenesis (research)

STEP 1 STEP 2 STEP 3 è STEP 4

Type è Major anatomical involvement

è Severity è   Detailed regional distribution and extent of ARTAG

Subpial è

MTL è

occasional or

numerous (focal/widespread)

è   Inf. Temporal Gy Hippocampus Amygdala Lobar è è   Frontal Parietal Occipital Lateral temporal Subcortical è è   Basal forebrain - - Brainstem è è   Mesencephalon Pons Medulla obl

Subependymal è

MTL è è   Temporal horn Lobar è è   LV: Frontal horn LV: Occipital horn Subcortical è è   LV: Caudate 3V: Thalamus Brainstem è è   Aq/Mesencephalon Aq/Pons Aq/Medulla obl.

Gray matter è

MTL è è   Inf. Temporal Gy Hippocampus Amygdala Lobar* è è   Frontal Parietal Occipital Lateral temporal Subcortical è è   Accumbens Caud/Put GP Basal forebrain Brainstem è è   Substantia nigra Pons Medulla obl.

White matter è

MTL è è   Inf. Temporal Gy Hippocampus Amygdala Lobar* è è   Frontal Parietal Occipital Lateral temporal Subcortical è è   Internal capsule Subinsular Pencil fibers Brainstem è è   Cerebral pedunculi Pyramids Midline

Perivascular è

MTL è è   Inf. Temporal Gy Hippocampus Amygdala Lobar è è   Frontal Parietal Occipital Lateral temporal Subcortical è è   Accumbens Caud/Put GP Basal forebrain Brainstem è è   Mesencephalon Pons Medulla obl.

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Figure'1.''''''''''

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Figure'2.'''

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Figure legends

Figure'3.''

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Figure'4.''''''''

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Figure 1. Comparison of tau (using AT8 antibody) immunoreactivities seen in primary

tauopathies with those observed in aging-related tau astrogliopathy (ARTAG).

Figure 2. Representative photomicrographs of ARTAG types.

Plump cytoplasmic tau immunoreactivity of astrocytes and tau-positive lining in subpial (a)

and subependymal (b) location. Perivascular-type: tau-immunoreactive astrocytic processes

arranged around vessels (c). White matter (WM)-type: Astrocytes in the subcortical white

matter with plump cytoplasmic immunoreactivity (d). Gray matter (GM)-type: Single-

appearing (e, f) or clusters (g, h) of astrocytes with fine granular tau immunoreactivity in the

processes without (e) or with (f) plump perinuclear cytoplasmic tau immunoreactivity.

The bar shown in “a” represents 30 µm for a, b, f; 50 µm for d, e, h; and 100 µm for c, g.

Figure 3. Representative images of different anatomical regions showing ARTAG.

a: Temporal cortex and white matter (WM); b: Dentate gyrus (gray matter-type cluster

enlarged in the right); c: Amygdala; d: Frontal cortex (gray matter-type single); e: Nucleus

accumbens (gray matter-type clusters and single forms); f: Substantia nigra; g and h: medulla

oblongata (IO: inferior olive; ML: medial lemniscus; n. XII: hypoglossal nucleus).

The bar shown in “a” represents 150 µm for a, b; 100 µm for the right inset in b, and c-h.

Figure 4. Summary of the concept of ARTAG.

Four distinct astroglial tau pathologies are seen in primary tauopathies: tufted astrocytes (TA),

astrocytic plaques (AP), globular astroglial inclusions (GAI), and ramified astrocytes (RA).

Rarely there may be slight overlap of these morphologies but predominance of a type is

significantly associated with one of the specific primary tauopathies. ARTAG is characterized

by two different morphologies: thorn-shaped astrocytes (TSA) and fine granular

immunoreactivity in astrocytic processes (granular/fuzzy astrocytes: GFA); these are seen in

the subpial (SP), subependymal (SE), perivascular (PV) areas, and in the white (WM) and

gray matter (GM). TSA and GFA may be present in the same brain together. Other

neurodegenerative diseases (NDDs) may coexist with ARTAG or with primary tauopathies.